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Open Access Research article

Identification and targeted disruption of the mouse gene encoding ESG1 (PH34/ECAT2/DPPA5)

Hisayuki Amano13, Ken Itakura1, Masayoshi Maruyama1, Tomoko Ichisaka12, Masato Nakagawa1 and Shinya Yamanaka12*

Author Affiliations

1 Department of Stem Cell Biology, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan

2 CREST, Japan Science and Technology Cooperation, Kawaguchi, Japan

3 Department of Molecular Biology and Biochemistry, Osaka University Graduate School of Medicine/Faculty of Medicine, Suita, Japan

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BMC Developmental Biology 2006, 6:11  doi:10.1186/1471-213X-6-11

Published: 28 February 2006

Abstract

Background

Embryonic stem cell-specific gene (ESG) 1, which encodes a KH-domain containing protein, is specifically expressed in early embryos, germ cells, and embryonic stem (ES) cells. Previous studies identified genomic clones containing the mouse ESG1 gene and five pseudogenes. However, their chromosomal localizations or physiological functions have not been determined.

Results

A Blast search of mouse genomic databases failed to locate the ESG1 gene. We identified several bacterial artificial clones containing the mouse ESG1 gene and an additional ESG1-like sequence with a similar gene structure from chromosome 9. The ESG1-like sequence contained a multiple critical mutations, indicating that it was a duplicated pseudogene. The 5' flanking region of the ESG1 gene, but not that of the pseudogene, exhibited strong enhancer and promoter activity in undifferentiated ES cells by luciferase reporter assay. To study the physiological functions of the ESG1 gene, we replaced this sequence in ES cells with a β-geo cassette by homologous recombination. Despite specific expression in early embryos and germ cells, ESG1-/- mice developed normally and were fertile. We also generated ESG1-/- ES cells both by a second independent homologous recombination and directly from blastocysts derived from heterozygous intercrosses. Northern blot and western blot analyses confirmed the absence of ESG1 in these cells. These ES cells demonstrated normal morphology, proliferation, and differentiation.

Conclusion

The mouse ESG1 gene, together with a duplicated pseudogene, is located on chromosome 9. Despite its specific expression in pluripotent cells and germ cells, ESG1 is dispensable for self-renewal of ES cells and establishment of germcells.